The central premise of this resubmitted, competitive renewal R01 application is to understand the biology of disease in Spinocerebellar Ataxia Type 3 (SCA3), the most common dominant ataxia worldwide. SCA3 is caused by abnormal CAG triplet repeat expansion in the gene ATXN3, which translates into a long polyglutamine (polyQ) tract in the deubiquitinase ataxin-3. How SCA3 occurs remains unresolved; this disease is uniformly fatal in patients. SCA3 belongs to the family of polyQ diseases, which includes another eight disorders. In each disease, the primary culprit of neuronal toxicity is the glutamine repeat within the primary sequence of its causative protein. PolyQ expansion beyond normal ranges leads to protein misfolding, neuronal dysfunction and death. There is clear evidence that the manner in which polyQ toxicity presents? CNS areas impacted and cellular processes perturbed?is regulated by peptide regions outside of the polyQ tract, what is commonly referred to as protein context. This fundamental aspect of polyQ degeneration is underscored by the fact that although a similar mutation causes each disease, the symptoms presented and the nervous system areas impacted differ among them. To understand the biology of polyQ diseases, we need to comprehend how the protein areas around each polyQ region control and modulate the toxicity of the expanded repeat. Our focus in this competitive renewal application is to systematically understand the role of protein context in SCA3. Our over-arching hypothesis is that pathogenicity conferred onto ataxin-3 by abnormal polyQ tract expansion is controlled by specific binding partners at this protein's non-polyQ domains. Our work during the prior cycle of this R01 award provided key clues about the role of protein context in SCA3. We found that two of its non-polyQ regions exert significant modulatory effect on degeneration caused by ataxin-3 in vivo. One region controls the interaction of ataxin-3 with the proteasome-associated protein Rad23; the other is used by the SCA3 protein to directly bind to the AAA ATPase VCP/p97. Each interaction has a specific and clear effect on the toxicity of pathogenic ataxin-3. In this cycle, we seek to expand on our findings and to propel our work through novel models and approaches. We propose a systematic and interdisciplinary set of studies to define the role of non-polyQ domains on the aggregation, toxicity and cellular properties of pathogenic ataxin-3 in vivo, while also investigating more broadly the cellular response to this toxic protein. We are confident that our work will add much needed perspective to the role of protein context in SCA3 from complementary and progressively wider angles, establish novel tools to study this disease, and provide new conceptual advances that will likely inform the biology of other proteotoxic disorders.